title: Molecular mechanism of an oncogenic mutation that alters membrane targeting: Glu17Lys modifies the PIP lipid specificity of the AKT1 PH domain.
abstract: The protein kinase AKT1 regulates multiple signaling pathways essential for cell function. Its N-terminal PH domain (AKT1 PH) binds the rare signaling phospholipid phosphatidylinositol 3,4,5-trisphosphate [PI(3,4,5)P(3)], resulting in plasma membrane targeting and phosphoactivation of AKT1 by a membrane-bound kinase. Recently, it was discovered that the Glu17Lys mutation in the AKT1 PH domain is associated with multiple human cancers. This mutation constitutively targets the AKT1 PH domain to the plasma membrane by an unknown mechanism, thereby promoting constitutive AKT1 activation and oncogenesis. To elucidate the molecular mechanism underlying constitutive plasma membrane targeting, this work compares the membrane docking reactions of the isolated wild-type and E17K AKT1 PH domains. In vitro studies reveal that the E17K mutation dramatically increases the affinity for the constitutive plasma membrane lipid PI(4,5)P(2). The resulting PI(4,5)P(2) equilibrium affinity is indistinguishable from that of the standard PI(4,5)P(2) sensor, PLCdelta1 PH domain. Kinetic studies indicate that the effects of E17K on PIP lipid binding arise largely from electrostatic modulation of the dissociation rate. Membrane targeting analysis in live cells confirms that the constitutive targeting of E17K AKT1 PH to plasma membrane, like PLCdelta1 PH, stems from PI(4,5)P(2) binding. Overall, the evidence indicates that the molecular mechanism underlying E17K oncogenesis is a broadened target lipid selectivity that allows high-affinity binding to PI(4,5)P(2). Moreover, the findings strongly implicate the native Glu17 side chain as a key element of PIP lipid specificity in the wild-type AKT1 PH domain. Other PH domains may employ an analogous anionic residue to control PIP specificity.
authors.affiliation: Department of Chemistry and Biochemistry and Molecular Biophysics Program, University of Colorado, Boulder, Colorado 80309-0215, USA.
authors.forename: Kyle E
authors.initials: KE
authors.lastname: Landgraf
authors.forename: Carissa
authors.initials: C
authors.lastname: Pilling
authors.forename: Joseph J
authors.initials: JJ
authors.lastname: Falke
mesh_terms: D016475:3T3 Cells; D000818:Animals; D002417:Cattle; D002462:Cell Membrane / Q000378:metabolism*; D002470:Cell Survival; D018698:Glutamic Acid / Q000235:genetics; D006801:Humans; D007700:Kinetics; D008239:Lysine / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008856:Microscopy, Fluorescence; D009154:Mutation; D009369:Neoplasms / Q000235:genetics*; D009857:Oncogenes / Q000235:genetics*; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000378:metabolism; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism*; D017434:Protein Structure, Tertiary / Q000235:genetics; D051057:Proto-Oncogene Proteins c-akt / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D013379:Substrate Specificity
chemical_list: D019269:Phosphatidylinositol 4,5-Diphosphate; D018129:Phosphatidylinositol Phosphates; C060974:phosphatidylinositol 3,4,5-triphosphate; D018698:Glutamic Acid; D051057:Proto-Oncogene Proteins c-akt; D008239:Lysine